Project Summary ? Overall The three Projects in this P01 application, all strongly supported by the P01 Core, address overlapping research areas related to the central goals of the grant of characterizing pathways controlling cancer at the level of gene regulation. With the advent of powerful technologies to examine gene expression at the single cell level, it is becoming apparent that transcriptional states are much more dynamic and heterogeneous than previously believed. This is true of normal cells and, even more so, in cancer cells. The three Projects in this P01 address this question from a variety of perspectives and with complementary experimental systems. The product of this research is expected to be a greater understanding of how transcriptional heterogeneity governs the plasticity of cellular states, including differentiation states, as cells progress from normalcy to malignancy. In Project 1, the Jacks laboratory will explore this question in the context of tumor progression in mouse models of lung adenocarcinoma. These studies will examine how such heterogeneity evolves over time, what controls the observed changes and transcriptional networks as well as how immune responses to cancer affect these processes. The theme of transcriptional heterogeneity will also be explored by the Lees laboratory in Project 3 using the same model systems. Project 3 will focus on the effects of mutation of the PRC1 component Bmi1 in these processes. As part of Project 2, the Sharp laboratory will explore the role of miRNAs in the regulation of gene expression, with a focus on how miRNAs function to control patterns of gene expression at the single cell level and establish distinct cellular states. This research will inform the analysis of data generated in Projects 1 and 3. Dr. Aviv Regev (Broad Institute and MIT) is an important collaborator on the research related to this theme. The expert bioinformatics support provided by the P01 Core is also an essential component of this research effort. Project 2 will also explore novel models of gene expression involving gel-sol transitions at transcription start sites and enhancer element, which could help explain the dynamic nature of gene expression as well as other aspects of transcriptional control. A second theme of the Program is the examination of the development of cancer stem-like and niche-like cells as well as other effects on cellular differentiation during tumor progression. This theme will be pursued by both Projects 1 and 3. In addition to other aspects, the P01 Core will be essential for studies examining the relevance of these finding to human cancer. The final theme concerns the examination of the cellular and molecular effects of inhibition of the epigenetic regulator PRMT5, which functions as an argenine dimethylase of Sm proteins involved in mRNA splicing. Projects 2 and 3 will collaborate on the study of the biology and biochemistry of PRMT5 inhibition with the ultimate goal of developing new anti-cancer agents for the treatment of human cancers, including glioblastoma and lung cancer.